Sample drying device as well as mass spectrometer and mass spectrometry system therewith

ABSTRACT

A channel ( 103 ) is formed in a substrate ( 101 ) and a drying area ( 107 ) comprising a plurality of pillars ( 105 ) is formed in one end of the channel ( 103 ). A cover ( 109 ) is formed over the channel ( 103 ), except the area above the drying area ( 107 ). When a sample is introduced into the channel ( 103 ), it is guided to the drying area ( 107 ) by capillary phenomenon. The drying area ( 107 ) is heated by a heater ( 111 ) to evaporate the solvent for concentrating and drying the solute.

BACKGROUND OF THE INVENTION

1. Field of the Invention

This invention relates to a sample drying device as well as a massspectrometer and a mass spectrometry system therewith.

2. Description of the Related Art

Microchips capable of separating a protein or nucleic acid have beenintensely investigated and developed (Patent document 1). On such amicrochip, there is formed a feature such as a micro-channel forseparation by fine processing, whereby an extremely small amount ofsample can be introduced into the microchip for separation.

However, in a separation process using a conventional microchip, acomponent separated is obtained as a solution or dispersion, so that inaddition to the microchip, a drying equipment is required for finallyproviding a dried material.

Analysis of the separated component is generally conducted by massspectrometry. For example, analysis using a MALDI-TOFMS (Matrix-AssistedLaser Desorption Ionization-Time of Flight Mass Spectrometer) has beensuggested as a method for efficiently ionizing a polymer compound formass spectrometry, and has been applied to proteomics analysis (Patentdocument 2).

However, when a polymer compound analyzed by mass spectrometry is abiological component such as a protein, a nucleic acid or apolysaccharide, a target component must be isolated from the biologicalsample in advance. For example, when analyzing a sample comprisingmultiple components, the sample is purified and then subjected to, forexample, two-dimensional electrophoresis for separating individualcomponents; each component is collected from each spot separated; andthen the collected component is used to prepare a sample for massspectrometry. Thus, a separation and a sample preparation processes mustbe separately conducted, leading to a cumbersome procedure.

In a MALDI-TOFMS, a measurement sample is prepared by blending a samplesolution with a matrix solution and adding dropwise the mixture to ametal-plate surface using an appropriate tool such as a micropipettewhen using an ion-generation promoting material called a matrix. Withouta matrix, a sample solution is applied dropwise to a plate in a similarmanner.

FIG. 6 illustrates a conventional process for preparing a sample forMALDI-TOFMS measurement. FIGS. 6(a) and 6(b) are a cross-sectional viewand a plan view, respectively, showing a sample solution 131 dropped onthe surface of a drying substrate 133. As shown in FIG. 6(b), themaximum width of the dropped sample solution 131 is significantly largerthan the maximum spot size 135 of a laser beam. As a result, a sampleconcentration per a unit area is lower and thus, a relatively largeramount of sample is required. The procedure is, therefore, not always asample preparation process suitable for analyzing a trace amount ofsample such as a biological component.

Furthermore, a sheet of drying substrate 133 is used for a plurality ofsamples in a conventional method. Thus, a drying process is needed foreach sample.

-   Patent Document 1: Japanese Laid-Open Patent Publication No.    2002-207031-   Patent Document 2: Japanese Laid-open Patent Publication No.    1998-90226

SUMMARY OF THE INVENTION

As described above, a drying device has been needed, which canefficiently concentrate and dry a small amount of sample such as abiological sample. In particular, there has been needed a drying devicewhich can efficiently dry a collected sample for mass spectrometry.

In view of the above situation, an objective of this invention is toprovide a small sample drying device capable of conveniently andefficiently concentrating and drying a sample, particularly a sampledrying device capable of continuously and efficiently drying a componentprepared by processing, for example, separation and purification, abiological sample.

Another objective of this invention is to provide a sample drying devicefor mass spectrometry for efficiently concentrating and drying a sample.A further objective of this invention is to provide a mass spectrometerequipped with a drying device, which is used as a substrate for sampledrying and mass spectrometry.

According to this invention, there is provided a sample drying devicecomprising a channel for a sample flowing in the channel and a sampledrying area having an opening communicating with the channel, whereinthe sample drying area comprises a fine channel narrower than thechannel.

In the sample drying device according to this invention, the sampledrying area has a narrower channel and an opening, so that a sample inthe channel is quickly guided to the sample drying area by capillaryphenomenon. The sample introduced in the sample drying area is quicklydried. As the sample in the sample drying area is dried, a samplesolution in the channel is spontaneously and continuously fed to thesample drying area. Thus, the drying device of this invention can beeasily operated and can efficiently dry the sample.

In this invention, “fine channel(s)” may be formed as, for example,

(i) voids between multiple protrusions formed in the drying area orbetween filling members such as beads;

(ii) pores in a porous material disposed in the drying area; or

(iii) concaves formed in the channel wall.

The fine channel preferably communicates with an opening. Thus, a sampledrying channel from the channel through the fine channel to the openingcan be ensured, so that the sample can be stably dried.

According to this invention, there is also provided a sample dryingdevice comprising a main channel for a sample flowing in the mainchannel; a plurality of side channels branched from the main channel anda sample drying area communicating with the side channels, wherein thesample drying area has a fine channel narrower than the side channels.

In the sample drying device, the sample drying area is formed in theside chain branched from the main channel, so that the sample can bequickly dried. The side channel can be made narrower than the mainchannel to ensure guiding a liquid from the main channel to the sidechannel.

In the device having such a configuration, a sample can be separated,prepared and/or analyzed as appropriate in the main channel, thenintroduced into the side channel and finally dried in the sample dryingarea. For example, the sample contains multiple components and the mainchannel may comprise a separating portion to separate the components.Such a configuration may allow the individual components in the sampleto be introduced to a plurality of side channels for preparing driedmaterials of these components. Thus, a single sample drying device canreadily perform multiple processes, for which multiple devices have beenemployed.

The sample drying device of this invention may comprise a temperaturecontroller for controlling a temperature of the sample drying area.Thus, the sample drying area may be selectively heated to continuouslyand more efficiently dry the sample and introduce the sample from thechannel to the sample drying area during the sample drying.

In the sample drying device of this invention, the sample drying areamay comprise a plurality of protrusions separated each other. A voidbetween the protrusions becomes a fine channel, which can ensureintroduction of a liquid by capillarity to promote sample drying.

The sample drying device of this invention may have a configurationwhere the sample drying area may be filled with multiple particles. Sucha configuration may be easily formed by filling the channel with theparticles from an opening. Thus, a narrower channel may be convenientlyformed in the sample drying area.

Alternatively, the sample drying device of this invention may have aconfiguration where the sample drying area is filled with a porousmaterial. As used herein, the term “porous material” refers to astructure having a fine channel communicating with the outside in bothsides.

The sample drying device of this invention may have a configurationwhere the top of the sample drying area projects from the opening. Thus,a surface area of the side wall of the sample drying area may be furtherincreased to further promote drying.

The sample drying device of this invention may have a configurationwhere the sample drying area has a lid comprising a fine channelcommunicating with the outside of the sample drying device. The finechannel in the lid communicating with the outer atmosphere allows aliquid to be guided from the channel to the fine channel in the lid bycapillary phenomenon, resulting in efficient drying. Furthermore, sincea dried sample is deposited over the fine channel, a surface area of thedried sample can be controlled by adjusting a width of the fine channelin the lid.

The sample drying device of this invention may have a configurationwhere a metal film is formed on the surface of the drying area. Thus, itmay be suitable as an electrode for applying an external force to anionized sample when being used as a sample holder in a massspectrometer.

According to this invention, there is also provided a mass spectrometercomprising a sample drying area included in the sample drying device asa sample holder. Since the mass spectrometer of this invention comprisesthe sample drying area as the sample holder, the sample holder may beused as the sample drying device. Thus, a pretreatment before conductingmass spectrometry, that is, the steps of separation, purification,analysis and collection by drying of components in a sample to bemeasure, may be continuously conducted in the sample holder, resultingin improved operability. A surface area of the dried sample may beadjusting by the size of the opening over the sample drying area. Thus,the sample may be formed into a shape corresponding to a spot system ofa laser beam applied to the sample during mass spectrometry. It canincrease a sample concentration in a laser irradiation area, to improveaccuracy and sensitivity of the measurement. Even in a small amount ofsample, a measurement sample can be, therefore, efficiently prepared andanalyzed.

According to this invention, there is also provided a mass spectrometrysystem comprising separating unit separating components in a biologicalsample by their molecular sizes and properties; pretreatment unitpretreating the sample components separated by the separating unitincluding enzymatic digestion; drying unit drying the pretreated sample;and mass spectrometry unit conducting mass spectrometry for the driedsample, wherein the drying unit comprises the above sample dryingdevice. Herein, the biological sample may be obtained by extraction froman organism or by synthesis.

As described above, this invention may provide a small sample dryingdevice for readily and efficiently concentrating or drying a sample,which comprises a sample drying area having an opening and a finechannel narrower than a channel. This invention can also provide asample drying device for mass spectrometry for efficiently concentratingand drying a sample. This invention further provides a mass spectrometerequipped with a drying device used as a substrate for drying and massspectrometry of a sample.

BRIEF DESCRIPTION OF THE DRAWINGS

The above and other objects, features and advantages of the presentinvention will be more apparent from the following preferred embodimentsand the accompanying drawings, in which:

FIG. 1 shows a configuration of a drying device according to anembodiment of this invention;

FIG. 2 shows a configuration of a drying device according to anembodiment of this invention;

FIG. 3 shows a configuration of a drying device according to anembodiment of this invention;

FIG. 4 shows a configuration of a drying device according to anembodiment of this invention;

FIG. 5 schematically shows a configuration of a microchip according toan embodiment of this invention;

FIG. 6 illustrates a conventional method for preparing a sample for massspectrometry;

FIG. 7 is a process cross-sectional view illustrating a process formanufacturing a drying device according to an embodiment of thisinvention;

FIG. 8 is a process cross-sectional view illustrating a process formanufacturing a drying device according to an embodiment of thisinvention;

FIG. 9 is a process cross-sectional view illustrating a process formanufacturing a drying device according to an embodiment of thisinvention;

FIG. 10 illustrates a drying device according to an embodiment of thisinvention when it is filled with a liquid;

FIG. 11 illustrates a change in a sample liquid when it is heated by aheater in a drying device according to an embodiment of this invention;

FIG. 12 schematically shows a configuration of a mass spectrometer;

FIG. 13 is a block diagram of a mass spectrometry system comprising adrying device according to an embodiment of this invention;

FIG. 14 shows a configuration of a drying device according to anembodiment of this invention;

FIG. 15 schematically shows a configuration of a chip according to anembodiment of this invention;

FIG. 16 shows a configuration of a pillar disposed in a drying area in achip according to an embodiment of this invention;

FIG. 17 illustrates DNA exudation in a drying area in a chip accordingto Example; and

FIG. 18 illustrates a channel outlet in a drying area without a pillarin a chip according to Example.

DETAILED DESCRIPTION OF THE INVENTION

This invention will be described by means of an exemplary small dryingdevice for readily and efficiently concentrating and drying a sample.The drying device may be used as a sample holder for a mass spectrometersuch as a MALDI-TOFMS. In all of the drawings, analogous components aredesignated by the same symbol, whose description is omitted asappropriate.

First Embodiment

FIG. 1 shows a configuration of a drying device according to thisembodiment. FIGS. 1(a) and 1(b) are a plan view and a cross-sectionalview of a drying device 129, respectively.

In the drying device 129, substrate 101 comprises a channel 103, whichcomprises a drying area 107 having a plurality of pillars 105 in oneend. The channel 103 is covered by a cover 109, but not covered by thecover 109, that is, opened in the drying area 107. The bottom of thedrying area 107 can be temperature-controlled by a heater 111.

In the drying device 129, the drying area 107 comprises many pillars105. Thus, a sample liquid 141 can be charged such that it wets thewhole channel wall in the drying area 107. It will be described withreference to FIG. 10. FIG. 10 illustrates a drying device 129 filledwith a liquid. FIG. 10(a) illustrates a drying area 107 without pillars105 while FIG. 10(b) illustrates a configuration according to thisembodiment.

As shown in FIG. 10(a), without a pillar 105, a sample liquid 141 canwet only a part of the drying area 107 along a channel wall from thecover 109. On the other hand, in FIG. 10(b), there are provided pillars105, whereby the sample liquid 141 is introduced from a channel 103 to adrying area 107 by capillary phenomenon and thus fills the whole dryingarea 107. Thus, in the configuration in FIG. 10(b), the whole uppersurface of the drying area 107 can be covered by the sample liquid 141.Furthermore, the pillars 105 ensure an adequate specific surface area ina channel in the drying area 107. The drying device 129 having such aconfiguration exhibits a higher drying efficiency.

The drying device 129 has a configuration where a sample liquidintroduced from the channel 103 to the drying area 107 by capillaryphenomenon is heated by a heater 111 to efficiently evaporate a solvent.In the configuration shown in FIG. 10(b), the pillars 105 on the channel103 in the drying area 107 increases a specific surface area of thechannel in the sample drying area, that is, a surface area of the wallper a volume of the sample drying area, so that the sample can bequickly guided to the upper surface and be efficiently concentrated inthe drying area 107. Then, the sample components are precipitated on thesurface of the drying area 107 and dried. Since the sample liquid 141 iscontinuously fed from the channel 103 to the drying area 107, operationis simple. In contrast, in the configuration shown in FIG. 10(a), thesample liquid is in contact only with the bottom and the sides of thechannel 103, a heating efficiency is lower than that in theconfiguration in FIG. 10(b).

A temperature of heating the drying area 107 by the heater 111 may beappropriately selected, depending on some factors such as properties ofcomponents in the sample liquid to be dried; for example, 50° C. to 60°C. both inclusive. Alternatively, a drying rate of the sample liquid inthe drying area 107 may be 0.1 μL/min to 10 μL/min both inclusive, forexample, 1 μL/min.

In the drying device 129, the lid 119 may have any shape by which thesubstrate 101 can be covered such that at least part of the upper partof the drying area 107 is opened. Since the channel 103 can be sealed byproviding the cover 109, the sample liquid in the channel 103 can bemore efficiently guided into the drying area 107. Furthermore, the sizeof the opening can be adjusted to control a shape of a dried sample asdiscussed in the sixth embodiment later.

The substrate 101 is made of silicon. The silicon surface is preferablyoxidized. Thus, the substrate surface becomes hydrophilic, so that asample channel can be suitably formed. Alternatively, the substrate 101may be made of another material such as a glass including quartz and aplastic. Examples of a plastic include thermoplastic resins such assilicon resins, PMMA (polymethylmethacrylate), PET(polyethyleneterephthalate) and PC (polycarbonate) and thermosettingresins such as epoxy resins. Such a material can be easily shaped,resulting in reduction in a manufacturing cost for a drying device.

When using these materials, a metal film may be formed at least over thewhole surface of the drying area 107. A metal film formed on the surfacemakes the device electro-conductive. Thus, when a sample after drying isanalyzed by mass spectrometry such as MALDI-TOFMS as a whole dryingdevice 129, a mass spectrometer may be simplified because the dryingarea 107 can be used as an electrode in the mass spectrometer forapplying an electric potential. Furthermore, it can prevent thecomponent of the substrate 101 from being sublimed along with a sample,to improve accuracy and sensitivity in measurement.

The substrate 101 may be made of a metal. Using a metal, an electricpotential can be more stably applied by the drying area 107, when asample after drying is analyzed by MALDI-TOFMS as a whole drying device129.

The pillars 105 may be, for example, formed by, but not limited to,etching the substrate 101 in a predetermined pattern.

The pillars 105 in FIG. 1 is cylindrical, but they may be, in additionto a pillar or pseudo-pillar, a cone such as circular cone and ellipticcone; a prism such as trigonal prism and quadrangular prism; and pillarshaving another cross-sectional shape. When the pillar 105 has across-sectional shape other than a pseudo-circle, the pillar 105 mayhave an irregular side, resulting in further increasing a surface areaof the side and further improving a liquid absorbing force by capillaryphenomenon.

Alternatively, a slit having the cross-section in FIG. 1(a) may beemployed in place of the pillar 105. When using a slit, the pillar 105may have any of various shapes such as a striped protrusion. Again, whenusing a slit, the side of the slit may be irregular to further increasea surface area of the side.

In terms of the dimensions of the pillar 105, a width may be, forexample, about 5 nm to 100 μm. In FIG. 1, a height is substantiallyequal to the depth of the channel 103. Variation in a height of thepillar 105 will be described in the forth embodiment.

A distance between adjacent pillars 105 may be, for example, 5 nm to 10μm.

The cover 109 may be, for example, made of a material selected fromthose for the substrate 101. The material may or may not be the same asthat for the substrate 101.

Next, there will be described a process for manufacturing a dryingdevice 129. The channel 103 or the pillars 105 may be formed on thesubstrate 101 by, but not limited to, etching the substrate 101 into apredetermined pattern.

FIG. 7, FIG. 8 and FIG. 9 are process cross-sectional views illustratingan exemplary manufacturing process. In sub-figures in each figure, themiddle is a top view and the right and the left are cross-sectionalviews. In this process, the pillars 105 are formed by the use ofelectron beam lithography using a calixarene as a resist for fineprocessing. The following is an exemplary molecular structure of acalixarene. A calixarene is used as a resist for electron beam exposureand may be suitably used as a resist for nano processing.

Herein, a substrate 101 is a silicon substrate with an orientation of(100). First, as shown in FIG. 7(a), on the substrate 101 are formed asilicon oxide film 185 and a calixarene electron-beam negative resist183 in sequence. Thicknesses of the silicon oxide film 185 and thecalixarene electron-beam negative resist 183 are 40 nm and 55 nm,respectively. Then, an area to be pillars 105 is exposed to an electronbeam (EB). The product is developed with xylene and rinsed withisopropyl alcohol. By this step, the calixarene electron-beam negativeresist 183 is patterned as shown in FIG. 7(b).

Next, a positive photoresist 137 is applied to the whole surface (FIG.7(c)). Its thickness is 1.8 μm. Then, the product is developed by maskexposure such that the area to be the channels 103 is exposed (FIG.8(a)).

Then, the silicon oxide film 185 is RIE-etched using a mixed gas of CF₄and CHF₃ to a thickness of 40 nm after etching (FIG. 8(b)). Afterremoving the resist by organic washing with a solvent mixture ofacetone, an alcohol and water, the substrate is subjected to oxidationplasma treatment (FIG. 8(c)). Then, the substrate 101 is ECR-etchedusing HBr gas. A height of the step in the substrate 101 after etching,in other words, a height of the pillars 105, is 400 nm (FIG. 9(a)).Next, the substrate is wet etched with BHF-buffered hydrofluoric acid toremove the silicon oxide film (FIG. 9(b)). Thus, the channel 103 and thepillars 105 are formed on the substrate 101.

Herein, it is preferable to make the surface of the substrate 101hydrophilic after the step in FIG. 9(b). By making the surface of thesubstrate 101 hydrophilic, a sample liquid can be smoothly guided intothe channel 103 and the pillars 105. In particular, in the drying area107 where the channel is finer by the pillars 105, hydrophilization ofthe channel surface is preferable because it may promote introduction ofa sample liquid by capillary phenomenon to improve a drying efficiency.

After the step in FIG. 9(b), the substrate 101 is heated in a furnace toform a silicon thermal oxide film 187 (FIG. 9(c)). Herein, heatingconditions are selected such that a thickness of the oxide film becomes30 nm. Forming the silicon thermal oxide film 187 can eliminatedifficulty in introducing a liquid into a separating device. Then, acover 189 is electrostatically joined. After sealing, the drying device129 is formed (FIG. 9(d)).

A metal film may be formed on the surface of the substrate 101. Themetal film may be made of a material such as Ag, Au, Pt, Al and Ti. Itmay be deposited by, for example, vapor deposition or plating such aselectroless plating.

When using a plastic material for the substrate 101, a known methodsuitable for the type of the material for the substrate 101 may beemployed, including etching, press molding using a mold such as embossmolding, injection molding and photo-curing.

Again, when using a plastic material for the substrate 101, the surfaceof the substrate 101 is preferably hydrophilized. By hydrophilizing thesurface of the substrate 101, a sample liquid can be smoothly introducedinto the channel 103 and the pillars 105. In particular, in the dryingarea 107 where the channel 103 is finer by the pillars 105,hydrophilization of the surface of the channel 103 is preferable becauseit may promote introduction of a sample liquid 141 by capillaryphenomenon to improve a drying efficiency.

Surface treatment for hydrophilization may be, for example, conducted byapplying a coupling agent having a hydrophilic group to the side wall ofthe channel 103. A coupling agent having a hydrophilic group may be, forexample, a silane coupling agent having an amino group, morespecifically; N-β(aminoethyl)γ-aminopropylmethyldimethoxysilane,N-β(aminoethyl)γ-aminopropyltrimethoxysilane,N-β(aminoethyl)γ-aminopropyltriethoxysilane,γ-aminopropyltrimethoxysilane, γ-aminopropyltriethoxysilane andN-phenyl-γ-aminopropyltrimethoxysilane. These coupling agents may beapplied by an appropriate method such as spin coating, spraying, dippingand vapor deposition.

Again, in terms of FIG. 1, a heater 111 for controlling a temperature ofthe drying area 107 is provided on the bottom of the substrate 101 asshown in FIG. 1(b). By disposing the heater 111 such that the end of thedrying area 107 is selectively heated, a sample liquid can be surelyintroduced from the channel 103 to the drying area 107 so that a dryingefficiency in the drying area 107 can be further improved.

Heating of the drying area 107 is more preferably conducted in anintermittent manner. FIG. 11 illustrates a change in a sample liquid 141during heating the drying area 107 by the heater 111. As shown in FIG.11(a), the drying area 107 is filled with the sample liquid 141 and thenheated by the heater 111. Then, drying proceeds and the amount of thesample liquid in the drying area 107 is reduced as shown in FIG. 11(b).When the heater is stopped after drying proceeds to some extent, thedrying area 107 is refilled with the sample liquid (FIG. 11(a)). Then,the heater 111 is again operated to restart drying (FIG. 11(b)). Theprocedure may be repeated to conduct both drying and introduction of thesample liquid in a balanced manner, resulting in improvement in a dryingefficiency.

Second Embodiment

FIG. 2(b) shows a configuration of a drying device according to thisembodiment. The configuration in FIG. 2(b) is as described for thedrying device of the first embodiment, except that a water absorber 115is formed in the drying area 107. The water absorber 115 has a surfacehaving a relatively hydrophilic porous structure, and a sample solutionis introduced from the channel 103 to the water absorber 115 filling thedrying area 107 by capillary phenomenon.

The water absorber 115 may have any shape where a sample liquid can beintroduced from the channel 103 to the drying area 107 by capillaryphenomenon and evaporated on the surface. The water absorber 115 may be,for example, porous silicon or porous alumina with an etched concavestructure formed by lithography.

Third Embodiment

FIG. 2(c) shows a configuration of a drying device according to thisembodiment. The configuration in FIG. 2(c) is as described for thedrying device in the first embodiment, except that the drying area 107is filled with beads 117. The beads 117 are fine particles whose surfaceis relatively hydrophilic. A sample solution is introduced from thechannel 103 to the beads 117 filling the drying area 107 by capillaryphenomenon.

The configuration in FIG. 2(c) can be provided by forming the channel103 in the surface of the substrate 101 as described in the firstembodiment and then filling one end of the surface with the beads 117.Herein, since the upper part of the channel 103 is opened, theconfiguration can be easily provided, because, for example, the beads117 can be smoothly placed.

The beads 117 may be made of any material whose surface is relativelyhydrophilic. In case of a highly hydrophobic material, its surface maybe hydrophilized. Examples of the material include inorganic materialssuch as glasses and various organic and inorganic polymers. The beads117 may have any shape which, when being placed, allows a channel forwater to be ensured; for example, particles, needles or plates. Forexample, the beads 117 as spherical particles may have an averageparticle size of 10 nm to 20 μm both inclusive.

Alternatively, the drying area 107 may be filled with metal beads orsemiconductor beads. Thus, an electric potential can be more surelyapplied by the drying area 107, when a whole drying device 129 isanalyzed by mass spectrometry such as MALDI-TOFMS.

Next, there will be described a method for filling the beads 117 in thechannel 103. Before joining the cover 109, a mixture of the beads 117, abinder and water is fed into the channel 103. Herein, a damming member(not shown) is formed in the channel 103 to prevent the mixture fromflowing outside the area to be the drying area 107. Then, the mixturecan be evaporated to dryness to form the drying area 107.

A binder may be, for example, a sol containing a water-absorbing polymersuch as agarose gel and polyacrylamide gel. A sol containing such awater-absorbing polymer can be used to eliminate the need of dryingbecause of spontaneous gelation. Alternatively, the drying area 107 maybe formed by filling the channel 103 with a suspension of the beads 117in water without a binder and drying it under the atmosphere of drynitrogen gas or dry argon gas.

Fourth Embodiment

FIG. 3(c) shows a configuration of a drying device according to thisembodiment. The configuration of the drying device in FIG. 3(c) is asdescribed in the first embodiment, except that the pillars 105 protrudefrom the opening.

FIG. 3(a) shows a configuration where a height of the pillars 105 issmaller than the depth of the channel 103; and FIG. 3(b) shows aconfiguration where a height of the pillars 105 is substantially equalto the depth of the channel 103 as described in the first embodiment.Since a surface area of the pillars 105 increases in the order of FIG.3(a), FIG. 3(b) and FIG. 3(c), a drying efficiency in the drying area107 is improved. In the configuration in FIG. 3(c), a sample is guidedto the part above the upper surface of the channel 103 by capillaryphenomenon and therefore a dried sample is also deposited in the upperpart of the channel 103. Thus, a dried target component can be moreeasily collected. Since a sample is concentrated in a direction of theheight of the drying area 107, measurement can be more accuratelyconducted in mass spectrometry such as MALDI-TOFMS.

Fifth Embodiment

FIG. 2(a) shows a configuration of a drying device according to thisembodiment. The configuration in FIG. 2(a) is as described in the firstembodiment, except that holes 113 are formed in the drying area 107.While a target component is concentrated, dried and deposited above thebottom of the channel 103 in the first to the forth embodiments, theconfiguration in FIG. 2(a) is different in that a target component isconcentrated, dried and deposited at the height near the bottom of thechannel 103. In the configuration where the holes 113 are formed in thedrying area 107, a surface area of the channel in the drying area 107 isalso increased by the holes 113, allowing a sample liquid to beefficiently concentrated and dried.

The configuration in FIG. 2(a) can be provided as described in the firstembodiment, for example, by etching.

Although the holes 113 have a circular cross section in FIG. 2(a), itmay have another shape such as a polygon. Furthermore, the side of thehole 113 may be made convexoconcave to further increase a surface areaof the side of the hole 113 as described in the first embodiment and tofurther increase a liquid absorbing force by capillary decreasing.

The holes 113 may be a slit having the cross section in FIG. 2(a). Whenusing a slit, a surface area of the side maybe also further increased bymaking the slit side irregular.

The hole 113 may have, for example, a width of 10 nm to 20 μm bothinclusive and a depth of 10 nm to 20 μm both inclusive.

Sixth Embodiment

This embodiment relates to a drying device where a sample is dried usingan opening formed in the upper part of a channel as a fine channel todeposit a dried sample on the upper surface of a lid. FIG. 14 shows aconfiguration of a drying device according to this embodiment. FIG.14(a) is a top view of a drying device 143 and FIG. 14(b) is across-sectional view of the periphery of the drying area 107 in FIG.14(a). The drying device 143 comprises a lid 119 covering the wholesurface of the channel 103 including the drying area 107. In the lid119, an opening 121 is formed as a fine channel, through which thechannel 103 is communicated with the outside air. Thus, a liquid in thesample introduced from the channel 103 to the drying area 107 is guidedto the opening 121 by capillary phenomenon and then evaporated.

The lid 119 formed allows a dried sample 123 to be selectively depositednear the opening 121 in the upper surface of the lid 119. Furthermore,the size of the opening 121 can be adjusted to adjust a surface area ofthe dried sample 123. One opening 121 may be formed in the lid 119 asshown in FIG. 14, or alternatively a plurality of openings 121 may beformed.

When forming the opening 121 in the lid 119 and, for example, the dryingdevice 143 and the dried sample 123 are analyzed by MALDI-TOFMSmeasurement, the size of the dried sample 123 may be adjusted to besubstantially equal to the maximum spot size 135 of a laser beamdescribed above in FIG. 6. Thus, a concentration of the dried sample 123can be increased in the laser-beam irradiation site to improve accuracyand sensitivity in measurement.

In the drying device 143, the pillars 105 may be formed in the dryingarea 107 as described in the first embodiment, which is shown in FIG.4(a). Thus, the channel becomes finer in the drying area 107, so thatdrying can be more efficiently conducted and the dried sample 123 can bedeposited near the opening 121 in the upper surface of the lid 119 (FIG.4(b)).

Seventh Embodiment

This embodiment relates to a microchip comprising a plurality of thedrying devices 127 described in the first embodiment. FIG. 5schematically shows a configuration of the microchip according to thisembodiment.

The microchip in FIG. 5 comprises a main channel 125 and a plurality ofside channels 127 branched from the main channel 125 on a substrate (notshown). Each side channel 127 is communicated with a plurality of dryingdevices 129.

Using microchip in FIG. 5, a sample liquid containing multiplecomponents can be purified and separated into the components, which canbe finally concentrated, dried and collected in the drying device 129.

For example, when a current is applied to the main channel 125 and theside channels 127 are filled with a gel and the like to conductseparation similar to two-dimensional electrophoresis in the microchip,the system can be designed such that a drying device 129 can becommunicated with a site corresponding to a band for each componentseparated in the side channel 127, to independently collect eachcomponent from the sample.

Specifically, for separating water-soluble proteins in blood, aseparating device may be placed upstream of the main channel 125 toremove insoluble components. Furthermore, a separation mechanism whichcan remove low molecular weight components in a plasma by permeation isemployed to allow only high molecular weight fractions to remain in themain channel 125. The remaining high molecular weight fractions aretwo-dimensionally separated in the main channel 125 and the sidechannels 127 as described above, before introducing them into the dryingdevice 129. Herein, the drying device 129 can be placed in the mainchannel 125 upstream of the side channels 127 to concentrate the highmolecular weight fractions to some degree before separation and thus tofurther improve a separation efficiency.

Although the drying device 129 is used in FIG. 5, a drying device havinganother configuration according to this embodiment may be, of course,employed.

Eighth Embodiment

In this embodiment, the drying device 129 according to the firstembodiment is used as a substrate for MALDI-TOFMS. There will bedescribed, as an example, preparation and measurement of a proteinsample for MALDI-TOFMS using the drying device 129.

For obtaining detailed data of a protein to be measured by MALDI-TOFMS,its molecular weight must be reduced to about 1000 Da. Thus, aftermolecular weight reduction, the sample is mixed with a matrix solutionand dried in the drying device 129 to provide a dried sample.

When the target protein has an intramolecular disulfide bond, the sampleis subjected to reduction in a solvent such as acetonitrile containing areducing agent such as DTT (dithiothreitol). Thus, a next decompositionreaction can efficiently proceed. It is preferable that after reduction,a thiol group is protected by, for example, alkylation to preventre-oxidation.

Next, the reduced protein molecule is subjected to molecular weightreduction using a protein hydrolase such as trypsin. Since molecularweight reduction is conducted in a buffer such as a phosphate buffer,desalting and removal of the high molecular weight fraction, that is,trypsin, must be conducted after the reaction. The material obtained ismixed with a MALDI-TOFMS matrix and introduced from the channel 103 tothe drying area 107.

A temperature in the drying area 107 is controlled by the heater 111 forconcentrating and drying the sample to precipitate a mixture of thematrix and the decomposed protein in the upper part of the pillars 105.Herein, as described above in the first embodiment, on-off of the heater111 can be repeated for repeating drying and introduction of the samplesolution to efficiently conduct drying.

After drying, the sample as a whole drying device 129 is set in aMALDI-TOFMS apparatus. Then, while applying a voltage using the dryingdevice 129 as an electrode, for example, it is irradiated with anitrogen laser beam at 337 nm for MALDI-TOFMS analysis.

There will be briefly described a mass spectrometer used in thisembodiment. FIG. 12 schematically illustrates a configuration of themass spectrometer. In FIG. 12, the dried sample is set on a samplestage. Then, the dried sample is irradiated with a nitrogen gas laser ata wavelength of 337 nm in vacuo, to vaporize the dried sample togetherwith the matrix. By applying a voltage using the sample stage as anelectrode, the vaporized sample travels in the vacuum atmosphere anddetected by a detection unit comprising a reflector detector, areflector and a linear detector.

Therefore, after fully drying the liquid in the drying device 129, thedrying device 129 can be placed in a vacuum chamber in the MALDI-TOFMSapparatus and used as a sample stage for MALDI-TOFMS. Since a metal filmis formed on the surface of the drying area 107 and is connectable to anexternal power source, a potential can be applied to it as a samplestage.

Thus, using the drying device 129, the dried sample as the whole dryingdevice 129 can be used in MALDI-TOFMS. Furthermore, for example, asample separating device may be formed upstream of the channel 103 to beable to conduct extraction, drying and structural analysis of a targetcomponent on a single drying device 129. Such a drying device 129 may beuseful in, for example, proteome analysis.

Herein, since the drying device 129 is used as a chip for MALDI-TOFMS, astep of washing an electrode plate for each sample can be eliminated,resulting in improvement in operational convenience and in measurementaccuracy.

A MALDI-TOFMS matrix may be appropriately selected, depending on amaterial to be measured. Examples of a matrix which can be used includesinapic acid, α-CHCA (α-cyano-4-hydroxycinnamic acid), 2,5-DHB(2,5-dihydroxybenzoic acid), a mixture of 2,5-DHB and DHBs(5-methoxysalicylic acid), HABA (2-(4-hydroxyphenylazo)benzoic acid),3-HPA (3-hydroxypicolinic acid), dithranol, THAP(2,4,6-trihydroxyacetophenone), IAA (trans-3-indoleacrylic acid),picolinic acid and nicotinic acid.

This embodiment has been described in terms of the drying device 129described in the first embodiment, but drying devices in otherembodiments can be, of course, used.

Alternatively, a fine-structure in the upper surface of the drying area107 comprising the pillars 105, the holes 113, the water absorber 115 orthe beads 117 and so forth in any of the drying devices described in theabove embodiments may be adjusted to allow a sample to be moreefficiently ionized without a matrix. Such a configuration can eliminatethe need for mixing a protein solution with a matrix solution, so that,for example, each fraction collected in the seventh embodiment togetherwith the drying device 129 may be used for MALDI-TOFMS.

FIG. 13 is a block diagram of a mass spectrometry system comprising adrying device according to this embodiment. As shown in FIG. 13(a), thesystem comprises means to perform each step of; purification 1002 forremoving impurities in a sample 1001 to some degree; separation means1003 for removing unnecessary components 1004; pretreatment 1005 for aseparated sample; drying 1006 for a sample after pretreatment; andidentification 1007 by mass spectrometry.

Drying by the drying device in this embodiment corresponds to the dryingstep 1006, which is conducted on a microchip 1008. The step ofpurification 1002 may be conducted, for example, using a separatingportion for separating only giant components such as blood cells. Thestep of separation 1003 may be conducted by a procedure such astwo-dimensional electrophoresis, capillary electrophoresis and affinitychromatography and so on. In the step of pretreatment 1005, molecularweight reduction using, for example, trypsin described above and mixingwith a matrix are conducted.

Since the drying device according to this embodiment comprises achannel, the steps of purification 1002 to drying 1006 may be conductedon a piece of microchip 1008 as shown in FIG. 13(b). A sample may becontinuously processed on the microchip 1008 to efficiently and reliablyidentify a trace amount of component in a loss-reducing manner.

Thus, of the sample processing steps shown in FIG. 13, all or thoseappropriately selected can be conducted on the microchip 1008.

This invention has been described with reference to some embodiments. Itwill be understood by the skilled in the art that these embodiments areonly illustrative and that there may be many variations for acombination of the components and the manufacturing process, which areencompassed by the present invention.

EXAMPLE

In this example, a drying device comprising the pillars described abovewith reference to FIG. 1 was fabricated on a substrate and evaluated.FIG. 15 schematically shows the drying device. FIG. 15(a) is a top viewof the drying device and FIG. 15(b) is a cross-sectional view taken online A-A′ of FIG. 15(a).

In FIG. 15, a channel 202 is formed on a substrate 201 and a part of itsupper surface is covered by a glass lid 203. The part with the glass lid203 is upstream while that without the lid is downstream. A drying area204 is formed in an outlet area in the channel 202, in other words, thearea upstream and downstream of the end of the glass lid 203. The dryingarea 204 comprises columnar structures 205.

In this example, the channel 202 and the columnar structure 205 wereformed by the processing method described in the first embodiment.Silicon was used as a substrate. The channel 202 had a width of 80 μmand a depth of 400 nm.

FIG. 16 shows a scanning electron microgram of the columnar structure205 formed in the outlet area in the channel 202. In FIG. 16 and FIGS.17 and 18 described later, the lower direction from the paper isupstream and the upper direction is downstream. As shown in FIG. 16, thedrying area 204 of the drying device of this example comprises aplurality of strip-type columnar structures 205 with a width of 3 μmaligned with an equal pitch of about 1 μm in a longitudinal direction ofthe columnar structures 205 (a transverse direction in this figure), andmultiple rows of the columnar structures 205 are disposed with an equalpitch of 700 nm in a lateral direction of the columnar structures 205 (avertical direction in this figure). A height of the columnar structures205 is 400 nm.

The drying device manufactured in this example was used to continuouslyconduct drying and mass spectrometry of a DNA as described below. Thechannel 202 was filled with a solution containing a DNA (100 bp) stainedwith a fluorescent dye from the upstream of the channel 202. Then, theoutlet area in the channel 202 was observed by fluorescence microscopy.FIG. 17 shows a fluorescence microgram of the area near the columnarstructure 205 formed in the drying area 204 in the outlet area in thechannel 202. FIG. 17 shows that the DNA brightly highlighted by thefluorescence microscopy is exuded as a 60 μm band downstream of theglass lid 203. Thus, using the drying device of this example, the samplecould be stably introduced into the drying area 204 and easily dried asdescribed with reference to FIG. 10(b).

For comparison, a drying device without columnar structures 205 wasmanufactured in a similar manner. FIG. 18 shows a fluorescence microgramfor the device without columnar structures 205 in the outlet area in thechannel, in which DNA is not exuded outside of the glass lid 203. In thechip used in this example without columnar structures 205 where thedepth of the channel 202 is 400 nm, it can be seen that a wetting degreedescribed with reference to FIG. 10(a) is further reduced so that thedrying area 204 is not wetted even in the area from the edge of theglass lid 203 to the wall surface of the channel 202.

Then, the DNA dried using the drying device in FIG. 17 was analyzed bymass spectrometry. Specifically, the substrate 201 was sonicated on anultrasonic vibrator to fragmentate the DNA and then the solvent was airdried. Then, a several microliters of matrix was added dropwise to thedried DNA exuded in the outlet area in the channel 202 and the productwas analyzed by MALDI-TOFMS. As a result, the analysis results from theDNA could be obtained.

As described above, in this example, the drying area 204 comprising aplurality of columnar structures 205 at the end of the channel 202 whoseupper surface is at least partly opened was formed, so that the DNAcould be moved to the drying area 204 and then easily dried.Furthermore, the drying device could be used as a sample stage for amass spectrometer and mass spectrometry could be conducted withoutremoving the dried sample from the drying device.

1. A sample drying device comprising: a channel for a sample flowing insaid channel; and a sample drying area having an opening communicatingwith said channel, wherein said sample drying area comprises a finechannel narrower than said channel.
 2. A sample drying devicecomprising: a main channel for a sample flowing in said main channel; aplurality of side channels branched from said main channel; and a sampledrying area communicating with said side channels, wherein said sampledrying area has a fine channel narrower than said side channels.
 3. Thesample drying device as claimed in claim 2, wherein said sample containsmultiple components and said main channel comprises a separating portionto separate said components.
 4. The sample drying device as claimed inany of claims 1 to 3, wherein said sample drying area comprises aplurality of protrusions separated each other.
 5. The sample dryingdevice as claimed in claim 4, wherein said drying area has a shape sothat the top of said sample drying area projects from said opening. 6.The sample drying device as claimed in any of claims 1 to 3, whereinsaid sample drying area is filled with multiple particles.
 7. The sampledrying device as claimed in any of claims 1 to 3, wherein said sampledrying area is filled with a porous material.
 8. The sample dryingdevice as claimed in any of claims 1 to 3, wherein said sample dryingarea has a lid comprising a fine channel communicating with said outsideof said sample drying device.
 9. The sample drying device as claimed inany of claims 1 to 3, wherein said sample drying device comprises atemperature controller for controlling a temperature of said sampledrying area.
 10. A mass spectrometer comprising a sample drying areaincluded in said sample drying device as claimed in any of claims 1 to3, as a sample holder.
 11. A mass spectrometry system comprising:separating unit separating components in a biological sample by theirmolecular sizes and properties; pretreatment unit pretreating saidsample separated by said separating unit including enzymatic digestion;drying unit drying the pretreated sample components; and massspectrometry unit conducting mass spectrometry for the dried sample,where in said drying unit comprises said sample drying device as claimedin any of claims 1 to 3.